| Experiment Id | GSE181527 | Name | The Heterogeneous Nuclear Ribonucleoprotein U (HNRNPU) is the Safeguard of Neural Stem Cell Viability |
| Experiment Type | RNA-Seq | Study Type | WT vs. Mutant |
| Source | GEO | Curation Date | 2024-11-06 |
| description | To study Hnrnpu function during cortical development we preformed single-cell RNA sequencing (scRNA-seq) on mouse neutrosphere cultures derived from E13 ICR cortices, 24 hours following treatment with CRISPR/CAS9 and two sgRNA sequences targeting mouse Hnrnpu. In parallel, we analyzed bulk RNA-seq from dissected cortices of E13 embryos in which Hnrnpu conditional truncation in the telencephalon was driven by Emx1::Cre. Additionally, we analyzed MARS-Seq trancriptomal profiles of cortices of E13 embryos carrying homozygous heterozygous or wildtype alleles of Hnrnpu truncation (Emx1::Cre driver) in combination with homozygous or heterozygous conditional deletion of Tp53 For scRNA seq, neurospheres were prepared from E13 ICR cortices, cultured for two days in the presence of EGF (20ng/ml), and bFGF (20ng/ml). Neurospheres were dissociated using Accutase and electroporated with a DNA mixture containing 4ug of pX330 CRISPR/CAS9 Hnrnpu Guide A, 4ug of pX330 CRISPR/CAS9 Guide B, and , 1ug of pCAG::GFP. Cells were collected 24 hours post electroporation, dissociated using Accutase and passed through a 40 micron strainer. The 10x Genomics Chromium system was used to capture 5000 single cells. The sequencing library was generated using the 10x Genomics Single Cell 3' Solution kit and subjected to Illumina NextSeq 550. For Bulk RNA seq and MARS seq, embryos (E13) were collected, their cortices were dissected in cold PBS, flush frozen in liquid nitrogen and kept in -80°C. RNA of desired genotypes was prepared using RNeasy Mini kit, Quiagen. |
